The standard method for Legionella testing is Legionella culture using the established methodology defined by the International Organization for Standardization (ISO).

However, many labs that follow the ISO method perform the minimum requirements for detection. This explains the variation in test results and pricing among laboratories.

For example, some labs use only one type of growth media. Some labs don’t use pre-treatment techniques that inhibit other bacteria and allow Legionella detection. While other labs use only the minimum method for identification, which tells you that you have Legionella but not what kind.

Because of our commitment to preventing Legionnaires' disease, Special Pathogens Laboratory follows both ISO 11731 and 11731-2. We use multiple culture media with a variety of antimicrobials to inhibit bacteria and foster Legionella growth. We use not just one but two pretreatment techniques—acid and heat. We concentrate samples to improve recovery of bacteria in water. And we use an extensive combination of methodologies to provide a definitive identification of the Legionella that is isolated. These include: latex aglutination, DFA, (monovalent and polyvalent) and DNA sequencing.

Q: What is a safe level for Legionella?

A: To date there is no agreement on a level (concentration) of Legionella in water that experts can say is safe or unsafe. Some guidelines or international standards advise when to take action based upon concentrations, but these levels haven't been scientifically validated and at best are overly conservative and arbitrary.

The level of CFU/ml hasn't been shown to be predictive of risk for Legionnaires' disease because Legionella infections are caused by the perfect storm of several factors including:

the virulence of Legionella species

mode of transmission (route and ability of the bacteria to enter the lungs)

health of the individual.

To better understand why establishing a minimum/maximum "contaminant" level has been so elusive and problematic, let's break it down:

Not all Legionella species have been shown to cause human disease (see article below, Assessing Risk: Nonpathogenic Legionella Species, which looks at the threat caused by non-pneumophila species)

Concentration of Legionella recovered (CFU/mL) is variable due to sample collection, culture methods and site specific conditions. Reliance on concentration to estimate risk is problematic and that is why we recommend a presence/absence approach.

However, some public health agencies, notably the CDC, describe this lack of predictable definable risk for all species, concentrations and routes of transmission as "there is no safe level of Legionella." This is often interpreted to mean there is no known safe level of any Legionella species. "Zero-tolerance" during an outbreak investigation may be a prudent response in stopping an outbreak but not for routine operations. Here's why :

Applying zero to a naturally occurring bacteria commonly found in water systems and difficult to completely eliminate is impractical.

Getting to zero and staying there is nearly impossible because building water systems are complex and many Legionella species can be present at any given time.

Limiting Legionella (of any species or serogroup) to zero as an enforceable limit is scientifically unsubstantiated and would result in time-consuming and costly interventions, such as has already been seen with the recently implemented VA Legionella Directive 1061.

Q: Should equipment for patient care such as transesophageal echocardiogram (TEE) probes be rinsed with hospital tap water?

A: Healthcare personnel should be aware that tap water isn't sterile and can cause infection. Legionella and other waterborne pathogens can be transmitted to patients when:

Equipment is mixed rinsed with tap water

Tap water is used for oral care.

Because of these reasons we don't recommend rinsing TEE probes with tap water.

Q: How effective is chlorine in controlling Legionella?

A: In the toolbox of Legionella disinfection technologies, chlorine gives you short term gain with long term pain.

In emergency situations, hyperchlorinating can give you short term relief. However, when used as a long term disinfectant there isn't much to recommend. Legionella is more tolerant of chlorine; it survives intracellularly up to 50 ppm. Studies show that rapid recolonization/rebound occurs if the chlorination system fails or levels drop. Continuous chlorination causes extensive corrosion and facilities using chlorine often switch to other modalities (i.e., chlorine dioxide, copper-silver ionization, monochloramine) due to the significant maintenance cost and need for corrosion control (phosphate or silicate injection). What's more, there's inadequate penetration of the agent into biofilms in piping and elevated temperatures and water pH compromises efficacy (> PH 8.5). If that' s not enough, chlorine produces carcinogenic byproducts (Trihalomethanes and Haloacetic acids).

Q: What volume of water should be collected for Legionella testing?

A: You will see on the SPL's Chain of Custody form that the minimum volume of water required for Legionella testing is 120 ml.

In the lab, we handle cooling and process water differently than potable water due to the different concentrations of competing bacteria. Because potable water contains less bacteria than cooling and process water, we concentrate it by filtration.

Note: It isn't necessary to collect large volumes (1,000 ml) of water for Legionella testing when performing routine testing to assess risk or verifying the efficacy of a disinfection method.

Q: When collecting a sample for Legionella testing should I use swabs or water? Is one better than the other?

A: For routine testing, collection of the first draw hot water--filling the 250 ml sample bottle--is recommended. Swab samples can also be used, but sometimes a swab can be negative when the water is positive.

Swabs sample the biofilm lining the fixture or pipe and the immediate draw hot water captures the bacteria sloughing off into the water. When using swabs, remember to remove the aerator before swabbing a faucet to get access to the biofilm.

When conducting an investigation, related to a case of Legionnaires' disease, it's best to collect both swab and water as this maximizes the opportunity to recover Legionella. For more information see SPL's Legionella sample collection instructions.

Q: Is there a one-size-fits-all approach to assessing risk of illness from Legionella in water systems?

A: Assessing risk should include characterizing three elements:

probability of exposure

likelihood of illness after exposure (susceptibility)

presence of Legionella.

​Since these factors are not consistent among all water systems, there is no one-size-fits-all approach for assessing risk.

The best place to start an assessment is to determine if Legionella is present. Other factors are relevant only when the hazard (Legionella) is present. Many people dismiss this first step because of the mistaken belief that Legionella is everywhere, and therefore nothing can be learned by testing for this waterborne pathogen. Numerous studies have evaluated the prevalence of Legionella in hospitals and office buildings. Depending on the study, the probability of finding Legionella in a water distribution system can be as low as 20% or as high as 70%. Samples tested from cooling towers range between 10% and 50% positive for Legionella.

Whether you're determining whether there is a risk for exposure to Legionella from your water system or are attempting to control this hazard using water treatment, you need to know what type of Legionella is present. If Legionellapneumophila serogroup 1 is in your system, the risk of illness is higher.

Q: Can you get a false positive Legionella culture?

A: No. A positive Legionella culture (colonies of Legionella growing on a petri dish) means Legionella was in the water sample.

Standard laboratory culture method, the industry standard, remains the most reliable detection method to date. When compared to culture, other methods can fail to detect Legionella when it's there or show a positive reading when Legionella isn't there.

For this reason, all other tests are compared to standard laboratory culture. Unfortunately, some rapid tests, like Direct Fluorescent Antibody (DFA), the Polymerase Chain Reaction (PCR), and dip slide methods, can give a false positive or false negative compared to culture.

For example, DFA stains are known to react with bacteria other than Legionella giving a false positive test result. Specifically, the Legionella strain can react with bacteria such as Bacillus (soil bacteria).The test result was false positive because the test confused Bacillus for Legionella.

When the safety of your water is evaluated, you want a reliable test to detect Legionella. So, rely on the industry "gold standard" of laboratory culture methods for validated and evidence-based results.

Q: How can hospitals with a secondary disinfection technology still have cases or an outbreak?

A: Because water systems are dynamic, levels of disinfectant and Legionella are in constant flux. Preventing hospital acquired Legionnaires' disease depends on effective management of the disinfection system and monitoring both the biocide and Legionella.

Events like disruption of water flow caused by water main breaks, construction, and ward closures can impact the flow of disinfectant through the water supply. Many hospitals simply turn the water back on after renovations not realizing that during this period, Legionella may have re-colonized in the plumbing system.

Additionally, once disinfection has been stopped for whatever reason, Legionella can rebound rapidly. In the early 1990s, a Pittsburgh hospital switched off its chlorine system that resulted in illness and death among some patients in the transplant unit.